title: Molecular cloning and identification of the transcriptional regulatory domain of the goat neurokinin B gene TAC3.
abstract: Neurokinin B (NKB), encoded by TAC3, is thought to be an important accelerator of pulsatile gonadotropin-releasing hormone release. This study aimed to clarify the transcriptional regulatory mechanism of goat TAC3. First, we determined the full-length mRNA sequence of goat TAC3 from the hypothalamus to be 820 b, including a 381 b coding region, with the putative transcription start site located 143-b upstream of the start codon. The deduced amino acid sequence of NKB, which is produced from preproNKB, was completely conserved among goat, cattle, and human. Next, we cloned 5'-upstream region of goat TAC3 up to 3400 b from the translation initiation site, and this region was highly homologous with cattle TAC3 (89%). We used this goat TAC3 5'-upstream region to perform luciferase assays. We created a luciferase reporter vector containing DNA constructs from -2706, -1837, -834, -335, or -197 to +166 bp (the putative transcription start site was designated as +1) of goat TAC3 and these were transiently transfected into mouse hypothalamus-derived N7 cells and human neuroblastoma-derived SK-N-AS cells. The luciferase activity gradually increased with the deletion of the 5'-upstream region, suggesting that the transcriptional suppressive region is located between -2706 and -336 bp and that the core promoter exists downstream of -197 bp. Estradiol treatment did not lead to significant suppression of luciferase activity of any constructs, suggesting the existence of other factor(s) that regulate goat TAC3 transcription.
authors.affiliation: Laboratory of Animal Production Science, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Suetomi
authors.forename: Fuko
authors.initials: F
authors.lastname: Matsuda
authors.forename: Yoshihisa
authors.initials: Y
authors.lastname: Uenoyama
authors.forename: Kei-ichiro
authors.initials: K
authors.lastname: Maeda
authors.forename: Hiroko
authors.initials: H
authors.lastname: Tsukamura
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ohkura
mesh_terms: D024506:5' Flanking Region; D000595:Amino Acid Sequence; D000818:Animals; D017849:Animals, Inbred Strains; D001483:Base Sequence; D002460:Cell Line; D017124:Conserved Sequence; D004967:Estrogens / Q000494:pharmacology; D005260:Female; D005300:Fertility Agents, Female / Q000494:pharmacology; D017930:Genes, Reporter / Q000187:drug effects; D006041:Goats; D006801:Humans; D007564:Japan; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D015287:Neurokinin B / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017385:Sequence Homology; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D004967:Estrogens; D005300:Fertility Agents, Female; D011994:Recombinant Proteins; D015287:Neurokinin B